This invention relates, in part, to newly identified polynucleotides and polypeptides; variants and derivatives of the polynucleotides and polypeptides; processes for making the polynucleotides and the polypeptides, and their variants and derivatives; agonists and antagonists of the polypeptides; and uses of the polynucleotides, polypeptides, variants, derivatives, agonists and antagonists. In particular, in these and in other regards, the invention relates to polynucleotides and polypeptides of a mouse chemokine receptor, hereinafter referred to as xe2x80x9cmouse CC-CKR5xe2x80x9d.
This invention relates to newly identified polynucleotides, polypeptides encoded by such polynucleotides, the use of such polynucleotides and polypeptides, as well as the production of such polynucleotides and polypeptides. More particularly, the polypeptides of the present invention are chemokine receptors. The invention also relates to inhibiting or activating the action of such polypeptides.
Chemokines are small signaling proteins that can be divided into two subfamilies, CC-chemokines or CXC-chemokines, depending upon the relative position of the first two conserved cysteines. A number of chemokines, including interleukin-8, RANTES, MCP-1, MCP-2, MCP-3, GROxcex1, GROxcex2, GROxcex3, MIP-1xcex1, and MIP-1xcex2 have been described. See, e.g. Baggiolini et al., ADVANCES IN IMMUNOLOGY (Dixon, F.J., Ed.)Vol. 55, pages 97-179, Academic Press, New York 1994. Chemokines attract and stimulate specific subsets of leukocytes and thus are very important regulators in the physiology of acute and chronic inflammatory processes. Oppenheim et al., Annu. Rev. Immunol., 1991, 9:617-648. For example, RANTES is a chemoattractant for monocytes, memory T-cells and eosinophils and induces the release of histamine by basophils. MCP-1, which is released by smooth muscle cells in arteriosclerotic lesions, is believed to be a factor responsible for macrophage attraction leading to the progressive aggravation of these lesions.
Recent studies have demonstrated that the actions of both CC- and CXC-chemokines are mediated by subfamilies of G-protein coupled receptors. Several functional receptors for CC and CXC chemokines have been identified in humans. For example, two receptors for interleukin-8 have now been identified. Holmes et al., Science, 1991, 253:1278-1283 and Murphy, P.M. and Tiffany, H.L. Science, 1991, 253:1280-1283. The first, IL-8RA, binds interleukin-8 specifically, while the second, IL,-8RB, binds IL-8 and other CXC-chemokines such as GRO. Several CC-chemokine receptors have also been identified. A receptor designated CC-chemokine receptor 1 or CC-CKR1 which binds both RANTES and MIP-1xcex1 was identified by Neote et al., Cell, 1993, 72:415-425. A second CC chemokine receptor CC-CKR2 which binds to MCP-1 and MCP-3 has also been identified. Charo et al., Proc. Nat""l Acad. Sci. USA, 1994, 91:2752-2756; Yamagami et al., Biochem. Biophys. Res. Commun., 1994, 202:1156-1162 ; Franci et al., J. Immunol., 1995, 154:6511-6517. CC chemokine receptors, CC-CKR3 and CC-CKR4, have also been identified. See, Combadiere et al., J. Biol. Chem. 1995, 270:16491-16494; Correction J. Biol. Chem., 1995, 270:30235 and Power et al., J. Biol. Chem., 1995, 270:19495-19500, respectively.
Molecular cloning and functional expression of a human receptor, ChemR13, was recently reported. Samson et al. Biochemistry, 1996, 35:3362-3367. The gene encoding ChemR13 is physically linked with the CC-CKR2 receptor gene in the human genome. The ChemR13 receptor, stably transfected in CHO-K1 cells was found to be stimulated by MIP-1xcex1, MIP-1xcex2, and RANTES, while MCP-1, MCP-2, MCP-3, IL-8 and GROxcex1x had no effect. This new human CC-chemokine receptor has been designated CC-CKR5.
A chemokine like orphan 7TM receptor, referred to as fusin, was also recently identified which is capable of acting as co-factor for the entry of the HIV-1 virus into T cells. Feng et al., Science, 1996, 272:872. Human CC-CKR5 receptor has also been reported as a principle cofactor in the entry of macrophage-tropic HIV-1 viruses into cells. These viruses are believed to be the primary pathogenic strain in vivo. Deng et al., Nature, 1996, 381:661; Dragic et al., Nature, 1996, 381:667.
A mouse ortholog of the human CC-CKR5 receptor has now been identified. Accordingly, it is now possible to knock out the mouse CC-CKR5 receptor gene from a mouse strain, replacing the mouse gene with human CC-CKR5 receptor gene to create a mouse model for human immunodeficiency virus. This mouse model is also useful in studying the role of this chemokine receptor in T-cell mediated inflammation.
Toward these ends, and others, it is all object of the present invention to provide polypeptides, inter alia, that have been identified as novel mouse CC-CKR5 by homology between the amino acid sequence set out in FIG. 1 and the known amino acid sequence of human CC-CKR5.
It is a further object of the invention to provide polynucleotides that encode mouse CC-CKR5, particularly polynucleotides that encode the polypeptide herein designated mouse CC-CKR5.
In a particularly preferred embodiment of this aspect of the invention, the polynucleotide comprises the region encoding mouse CC-CKR5 in the sequence set out in FIG. 1.
In accordance with this aspect of the present invention, there is provided an isolated nucleic acid molecule encoding a mature polypeptide expressible from the mouse gene contained in ATCC Deposit No. 98170.
In accordance with this aspect of the invention, there are provided isolated nucleic acid molecules encoding mouse CC-CKR5, including mRNAs, genomic DNAs and fragments and, in further embodiments of this aspect of the invention, biologically, diagnostically, clinically or therapeutically useful variants, analogs or derivatives thereof, including fragments of the variants, analogs and derivatives.
In accordance with another aspect of the present invention, there are provided methods of generating a mouse model for screening for compounds which bind to and activate or inhibit activation of the human CC-CKR5 receptor.
In accordance with another object the invention, there are provided products, compositions, processes and methods that utilize the aforementioned polypeptides and polynucleotides for research, biological, clinical and therapeutic purposes, inter alia.
In accordance with certain preferred embodiments of this aspect of the invention, there are provided products, compositions and methods, inter alia, for, among other things: assessing human CC-CKR5 expression by determining CC-CKR5 polypeptides or CC-CKR5-encoding mRNA; to treat T-cell mediated inflammation and more specifically HIV-infection, in vivo by exposing mice expressing human CC-CKR5 to a CC-CKR5 polypeptide or polynucleotide to augment CC-CKR5 function or remediate CC-CKR5 dysfunction.
In accordance with still another embodiment of the present invention, there is provided a process of using such activating compounds to stimulate the CC-CKR5 receptor for the treatment of conditions related to the under-expression of this receptor.
In accordance with another aspect of the present invention, there is provided a process of using such inhibiting compounds for treating conditions associated with over-expression of the CC-CKR5 receptor.
In accordance with yet another aspect of the present invention, there is provided non-naturally occuring recombinant CC-CKR5 polypeptides which are fragments, consensus fragments and/or sequences having conservaitive amino acid substitutions of at least one domain of the mouse CC-CKR5 of the present invention, such that the receptor may bind human CC-CKR5 ligands, or which may also modulate, quantitatively or qualitatively, human CC-CKR5 ligand binding.
In accordance with still another aspect of the present invention, there are provided synthetic or recombinant mouse CC-CKR5 polypeptides, conservative substitution and derivatives thereof, antibodies thereto, anti-idiotype antibodies, compositions and methods that can be useful as potential modulators of human CC-CKR5 function, by binding to ligands or modulating ligand binding, due to their expected biological properties, which may be used in diagnostic, therapeutic and/or research applications.
It is still another object of the present invention to provide recombinant polypeptides which are designed to inhibit or mimic various mouse CC-CKR5 or fragments thereof, as receptor types and subtypes.
In accordance with certain preferred embodiments of this and other aspects of the invention, there are provided probes that hybridize to CC-CKR5 sequences.
In certain additional preferred embodiments of this aspect of the invention, there are provided antibodies against mouse CC-CKR5 polypeptides. In certain particularly preferred embodiments in this regard, the antibodies are highly selective for mouse and human CC-CKR5.
Other objects, features, advantages and aspects of the present invention will become apparent to those of skill in the art from the following description. It should be understood, however, that the following description and the specific examples, while indicating preferred embodiments of the invention, are given by way of illustration only. Various changes and modifications within the spirit and scope of the disclosed invention will become readily apparent to those skilled in the art from reading the following description and from reading the other parts of the present disclosure.